The invention relates to a biosensor comprising an electrically conductive substrate, with a first layer comprising Ruthenium Purple formed on the substrate, a second layer comprising polyaniline or a derivative thereof comprising one or more non-polar substituents formed on the first layer, and a third layer comprising one or more enzymes trapped within a matrix formed on the second layer. The biosensor is for use in the detection of analytes such as purines and derivatives thereof, particularly in the detection of hypoxanthine.

Biosensors contain an electrode with a biorecognition molecule such as enzymes, antibodies, organelles and whole cells to allow the specific identification of analytes such as metabolytes, ions, gases and organic vapours. Such biorecognition elements are typically attached to a transducer, such as an electrochemical, piezoelectric, optoelectronic, fibreoptic, thermistor, diode or surface acoustic device, to enable the binding of the analyte to the biorecognition element to be detected. Amperometric biosensors, for example, are reviewed in the article by Palmisano F., et al. (Fresenius J. Anal. Chem. (2000), Vol. 366, pages 586-601). A wide range of mediator compounds and mediator polymers have been used to enable the transfer of electrons from a suitable biorecognition element, such as an enzyme, to an electrode.
Since the early work by Clark and Lyons (Clark and Lyons, 1962) biosensors have evolved continuously to become a powerful analytical tool in many fields. Immobilization of active enzymes central to the operation of biosensors has been achieved in many different ways (Cass, 1990). However entrapment of enzymes in electropolymer matrices (Bartlett and Cooper, 1993) has the particular advantage of producing biosensors with very small and custom shaped sensing elements. These types of micro-biosensors are ideal for detecting neurotransmitter release in the central nervous system as they are minimally invasive.
Cosnier and coworkers developed pyrrole derivatives that are suitable for entrapping enzymes on microelectrodes, and used these methods to develop a variety of biosensors including sensors sensitive to glutamate (Poitry et al., 1997) and dopamine (Cosnier et al., 1997). Indeed, biosensors using biotinised derivatives are shown in U.S. Pat. No. 6,197,881B in the name of Cosnier. Although these are important signaling agents in the nervous system, the polymer films on these sensors were insufficiently robust to survive implantation into neural tissue (Poitry et al., 1997) and the sensors had mixed success at detecting release from biological tissue (Poitry et al., 1997; Cosnier et al., 1997). Nevertheless, the methods developed by Cosnier are of potentially very general utility and the inventors have previously adapted them to construct robust and sensitive microelectrode biosensors that permit, spatially localized and fast detection of purine release from the nervous system. The purines, ATP and adenosine, perform extremely important signaling functions in both the peripheral and central nervous system. Peripherally, they are involved in the control of smooth muscle contraction and are powerful vasodilators (McMillan et al., 1999). Centrally, their diverse roles include regulation of spinal pain pathways (Sawynok, 1998), neuroprotection during ischaemia (Dale et al., 2000), control of transmitter release (Brundege and Dunwiddie, 1997), regulation of spinal motor pattern generation (Dale, 1998; Dale and Gilday, 1996) and induction of sleep (Porkka-Heiskanen, 1999). Adenosine, in particular, does not conform to the conventional paradigm of chemical neurotransmission in the nervous system. Instead of being released directly like most transmitters, adenosine is usually produced in the extracellular space from previously released ATP through the actions of special enzymes collectively known as the ectonucleotidases (Zimmermann and Braun, 1999). The production of adenosine—its spatial domains and kinetics of accumulation—are central to its function in the nervous system and can therefore be expected to differ considerably from those of conventional neurotransmitters. HPLC analysis of collected superfusate has been used to study adenosine release, however this method has very limited time and spatial resolution (Pedata et al., 1993). New methods for directly measuring adenosine production would thus be of great value in understanding its contribution to neural functions.
Recently, detection of adenosine produced during physiological activity has been achieved with an 3-enzyme biosensor (mark-1) (Dale, 1998) that utilizes a microdialysis electrode (250 mm diameter) to trap the required enzymes behind a semi-permeable membrane. This sensor is the subject of WO 99/07877 and is sensitive and has successfully detected release of adenosine from Xenopus embryo spinal cord (Dale, 1998) during motor activity, and mammalian hippocampus during hypoxia (Dale et al., 2000; Pearson et al., 2001). However it is too large to implant into nervous tissue without causing considerable damage and subsequent tissue reaction that may confound and invalidate physiological measurements. Furthermore the large size of the sensing assembly (500 μm when used in its finally constructed format) introduces diffusional delays which slows sensor responsiveness. To increase the range of applications for this measurement technology and to enable resolution of fast production of adenosine and related purines much smaller and faster responding biosensors are thus required.
The ability to monitor of the levels of purines such as hypoxanthine is also of vital importance in medicine. For example, hypoxanthine can provide an indicator of a baby's oxygen levels during birth and acts as an early warning system for distress.
If a baby's brain is being starved of oxygen, midwives and doctors have a more accurate idea of when a caesarean section is necessary. The ability to monitor the hypoxanthine levels can therefore help to prevent unnecessary emergency caesarean sections, as doctors sometimes perform one if they fear the baby is suffering foetal hypoxia, or oxygen starvation, which can result in cerebral palsy or other biological defects for a child. A baby with more than 5 μM of hypoxanthine per litre of blood is at a severe risk of foetal hypoxia. A level of 15 μM or more could indicate major hypoxian and a heightened risk of brain damage.
At present, blood samples are taken to a laboratory for analysis, which causes a delay in obtaining the results. It would therefore be beneficial to have a biosensor which would give doctors in the delivery room almost instant data on whether an unborn child faced the risk of hypoxia.
One alternative electrode is discussed in WO 03/087801. This discloses the use of sugar-derivatised pyrroles on a platinum or platinum-alloy substrate, in combination with a second layer comprising an ampiphilic pyrrole. This was found to improve the resilience of the sensor.
An alternative arrangement of the biosensor is shown in WO 2004/048603, which is incorporated herein by reference in its entirety. This describes methods of producing sol-gels on electrically conductive substrates. This allows the entrapment of, for example, enzymes into a sol-gel matrix for the production of biosensors.
EP 0537761 discloses a biosensor comprising an electrode on a substrate of polyethylene tetraphthalate. An enzyme, such as xanthine oxidase, is attached to an electrode via an electron acceptor, such as potassium ferricyanide, which is also known as Prussian Blue (PB).
However, the use of PB as an electron acceptor provides problems, most notably due to its intolerance to the presence of Na+ ions in physiological fluids. Na+ ions are prominent in physiological fluids such as blood, which clearly renders PB less suitable as a biosensor material.
It is therefore also necessary to develop a biosensor that is tolerant to the presence of Na+ ions in physiological fluids and can still readily function in such environments.
A related compound to PB, Ruthenium Purple (RP), which has the formula KFeRu(CN)6 or Fe4[Ru(CN)6]3, has also been used in biosensors. However, the only RP modified biosensors which have been described in the literature to date are all related to glucose biosensors, and clay, zeolite or cinder is used to anchor the RP in those biosensor configurations (see e.g. Zen et al, Anal. Chem., 2003, 75, 2703-2709).
RP does not possess the intolerance to Na+ ions that is exhibited by PB. Although the presence of Na+ ions has a minor effect upon the potential, the RP still functions to an effective level as an electron acceptor. In similar environments, similar biosensors comprising PB simply cease functioning.
The above properties are illustrated in the electrochemistry. The electrochemistry of RP is largely unaffected in ordinary phosphate buffer solutions which contain significant concentrations of Na+ ions (see FIG. 2). In order to maintain the electrochemical activity of PB, it is known that cations which fit the PB lattice, such as K+ Rb+, Cs+ and NH4+, are required. Other smaller cations in a buffer solution, such as Na+ Mg2+, H+ and Ca2+, act as blocking cations, negatively affecting the electrochemistry of the PB which impacts upon the operational potential of the biosensor. Hence the use of Na+ ions, which are widely employed in bioanalysis, has a significant restrictive effect upon the use of PB in biosensors.
Similarly to PB and its other analogues, RP has both soluble and insoluble forms. However, RP is considerably more soluble in water than PB, and is extremely stable in its insoluble oxidised form. At the optimum operational potential of the RP modified biosensor (which is usually 0 or −50 mV vs Ag/AgCl), RP is in its soluble form, which is susceptible to leaching, resulting in a loss of activity. A decrease in response is almost always observed in such biosensors. The ability to stabilise the RP against such leaching and boost the response would therefore be desirable.
Therefore, in accordance with the present invention, there is provided a biosensor comprising:                (i) an electrically conductive substrate;        (ii) a first layer formed on the substrate, the first layer comprising Ruthenium Purple;        (iii) a second layer formed on the first layer, the second layer comprising polyaniline or a derivative thereof comprising one or more non-polar substituents; and        (iv) a third layer formed on the second layer, the third layer comprising one or more enzymes. Preferably the enzymes are trapped within a matrix, or adsorbed onto the second layer. Most preferably a matrix is used.
In order to overcome the leaching problem associated with RP biosensors, a modified RP formation method is employed in the present invention, which has not previously been described. The present invention preferably involves stabilising the RP layer by heating, electrochemically cycling it in a solution of RuCl3, and contacting it with a solution containing aniline or a derivative thereof. In this way, a membrane comprising polyaniline or a derivative thereof comprising one or more non-polar substituents is introduced into the RP modified biosensor.
FIG. 4 depicts the FIA response of hypoxanthine on an RP modified biosensor in the absence of any polyaniline. It can clearly be seen that there is a decrease in the response with time to the hypoxanthine. This is due to the leaching of the RP. FIG. 5 also clearly shows the impact of the leaching of the RP, as the cyclic voltammogram of the biosensor without the polyaniline membrane becomes significantly smaller upon subsequent cycles. This is in contrast to the cyclic voltammogram B in FIG. 5, which shows little or no leaching upon repeated cycles.
FIG. 6 further confirms this as the FIA response to hypoxanthine is consistent over time. The polyaniline substantially prevents the leaching of the RP in solution and also does not interfere with the electrochemistry.
Examples of polyaniline films in biosensors include Iwouoha E. I., et al. (Biosensors & Bioelectronics (1997), 12 (8): 749-761, Chois J. and Park S. M. (J. Electrochemical Soc. (2002), 149 (2): E26-E34) and Sun, et al. (J. Chem. Soc. Chem. Commun. (1990), 529-531).
Preferably at least a portion of the polyaniline is in its oxidised state. The presence of oxidised aniline appears to be particularly important in preventing leaching of the RP. The oxidised form of polyaniline is understood to be conductive.
Hence, preferably at least a portion of the polyaniline is oxidised. This may preferably be achieved, for example, by cycling the polyaniline layer with a positive voltage, for example, up to +1.3V to oxidise the polyaniline or derivative thereof.
While polyaniline is the preferred component of the membrane, other polyaniline derivatives comprising one or more non-polar substituents may also be used within the scope of the invention. By non-polar, we mean a functional group which possesses a low dipole moment value, and which is composed either of elements having nonpolar covalent bonds or polar covalent bonds that cancel each other out. Such substituents include, in particular, alkyl chains, preferably C1-C12 alkyl chains, and ethers, preferably containing C1-C12 alkyl chains. Any straight chain or branched alkyl chains are included, especially Me, Et, n-Pr, i-Pr, n-Bu, t-Bu, n-Pe and n-He.
Other polyaniline derivatives comprising one or more polar substituents, such as hydroxy or further amino groups (e.g. poly(aminophenols) or poly(diaminobenzenes)), or other substituted benzenes such as polyphenols or polypyrrole, do not function particularly well in the biosensors of the invention, as they either interfere in the electrochemistry or fail to prevent leaching.
Although RP modified biosensors are known in the art, they have not been employed in the detection of chemicals such as purines and derivatives thereof, such as hypoxanthine, ATP, adenosine and/or inosine. The prevention of leaching in microelectrodes is particularly important.
Preferably, the electrically conductive substrate comprises platinum or a platinum alloy, gold or a gold alloy, or carbon. More preferably, however, the substrate is platinum or a platinum-iridium alloy, such as containing a 90:10 ratio of platinum:iridium (weight:weight), or gold.
Preferably, the one or more biological materials in the matrix layer are enzymes. Preferably, the enzymes are independently selected from xanthine oxidase, glucose oxidase, lactate oxidase, cholesterol oxidase, galactose oxidase, glutamate oxidase, horse radish peroxidase, polyphenol oxidase, D-fructose dehydrogenase, L-glutamate dehydrogenase, alcohol dehydrogenase (such as methanol dehydrogenase), urease, uricase, lactate dehydrogenase, glutamic pyruvic transaminase, creatinase, sarcosine oxidase, glutaminase, nucleoside phosphorylase, ascorbate oxidase, cytochrome C oxidase, adenosine deaminase, D- or L-amino acid oxidase, tyrosinase, catalase, phosphoenolpyruvate kinase, glycerol kinase, glycerol-3-phosphate oxidase, phosphocreatine kinase, acetylcholine esterase, choline oxidase and/or cholinedehydrogenase. Other enzymes known in the art may also be used. The enzymes may be used separately or two or more together in the form of a cascade to measure one or more different substrates. Typically, such electrodes use an oxidoreductase which is capable of transferring an electron, via the RP, onto the electrode.
Preferably, the analyte is selected from the following analytes, which are given below, together with the preferred combination of enzymes used for their detection:
Glucose (glucose oxidase), lactate (lactate oxidase), cholesterol (cholesterol oxidase), galactose (galactose oxidase), glutamate (glutamate oxidase), hypoxanthine (xanthine oxidase), hydrogen peroxide (horse radish peroxidase), fructose (D-fructose dehydrogenase), glutamate (L-glutamate dehydrogenase), ethanol (alcohol dehydrogenase), methanol (methanol dehydrogenase), urea (urease), uric acid (uricase and horse radish peroxidase), lactate (L-lactate dehydrogenase and glutamic pyruvic transaminase), creatine (creatininase and creatinase and sarcosine oxidase), glutamine and glutamate (glutaminase or glutamate oxidase), adenosine (adenosine deaminase), ATP (glycol kinase or glycol-1,3-phosphate oxidase), and inosine (nucleoside phosphorylase).
Most preferably, the analyte is hypoxanthine, ATP, adenosine and/or inosine.
The ability to induce several enzymes at the same time improves the speed of production of the biosensor and the response obtainable by such biosensors.
Other enzymes include RNase, DNase, nuclease, ribonuclease and catalase. Active proteins, such as haemoglobin, myoglobin, collagen or tubulin may be added. Antibodies or fragments of antibodies, such as IgG, IgM or Fab or F(ab1)2 fragments may also be incorporated.
Most preferably, the biosensors contain an oxidoreductase enzyme, such as xanthine oxidase. The biosensor may additionally comprise nucleoside phosphorylase and/or adenosine deaminase. The enzyme may be glycerol kinase and glycerol kinase and glycerol-3-phosphate oxidase.
Xanthine oxidase may be used, for example, to detect hypoxanthine. If xanthine oxidase is used together with nucleoside phosphorylase, then the electrode may be used to detect xanthine, inosine and hypoxanthine. The addition of adenosine deaminase to the enzymes converts purines, such as adenosine, into inosine, therefore allowing the detection of purines such as adenosine by the sensor. Inosine, in turn, may be converted into hypoxanthine by nucleoside phosphorylase.
Finally, in the chain hypoxanthine is converted into uric acid and hydrogen peroxide by xanthine oxidase. It is the hydrogen peroxide that is detected by the substrate.
Using all three enzymes allows the detection of purines, such as adenosine to be detected, as well as inosine, hypoxanthine and xanthine. Using only nucleoside phosphorylase and xanthine oxidase means that inosine, hypoxanthine and xanthine may be detected. Alternatively, using only xanthine oxidase, hypoxanthine may be detected.
Preferably, an excess of xanthine oxidase compared with nucleoside phosphorylase is used. Preferably, approximately equal amounts of nucleoside phosphorylase and adenosine deaminase are used.
Most preferably the ratio of adenosine deaminase:nucleoside phosphorylase:xanthine oxidase is approximately 1:1:5, based on units of activity. This ratio has been found to be the optimal ratio for this sort of electrode.
ATP may be detected with glycerol kinase and glycerol-1,3-phosphate oxidase. Enzymatic amplification can be achieved by including creatine kinase as well.
Although purine biosensors are exemplified here, use of other enzymes and enzyme cascades of two or more enzymes may be used.
In the biosensor of the invention, hydrogen peroxide is preferably detected at the electrode as a product of the bio-reaction. The response correlates with the concentration of the substance to be detected. The RP, in its role as an electrochemical mediator, eliminates the overpotential of detecting the hydrogen peroxide directly on the surface of the biosensor electrode, which can cause other biochemical reagents (such as ascorbic acid or dopamine) to be co-oxidised, resulting in interferences in the results obtained.
The matrix may be any suitable matrix known in the art for entrapping biological materials for use in biosensors. Such matrices are discussed in, for example, Palmisano F. (2000) supra and WO 03/08721.
Enzymes may also be adsorbed onto the surface, for example, using glutaraldehyde to cross link the protein to a layer of bovine serum albumin (BSA). Alternatively, specific linkers known in the art, such as biotin, carboxy or amine groups, may be used to attach the enzymes.
In the present invention, RP is electrodeposited onto an electrode surface, and a matrix layer containing biological materials is then grown on the layer of RP. The RP is substantially unaffected by this. The matrix layer may be grown according to the method detailed in WO 2004/048603, incorporated herein by reference in its entirety.
A negative potential is preferred to grow the optimum matrix layer on RP. The layer can also be grown galvanostatically or at predetermined cathodic currents.
Preferably, the matrix layer is a sol-gel layer. However, the matrix layer can also comprise a hydrogel, polyvinyl, polyacrylamide, polypyrrole, and sugar-derivatised polypyrroles.
By sol-gel we mean a colloidal suspension of sol particles that is gelled to form a solid. A sol is made of colloidal particles which are, prior to gelling, dispersed in a fluid such as liquid. Typically, colloidal particles have a size of 1 nm to 100 nm diameter.
The sol-gel layer preferably comprises a sol of a silicon-containing compound, such as a silicate or a silane (such as an alkoxysilane), alumina, a ceramic oxide sol or a colloidal metal hydroxide.
U.S. Pat. No. 6,303,290 discloses a ceramic oxide colloidal sol which may be mixed with an acidified salt solution. Adding hydroxide to increase the pH causes the formation of a sol-gel. Using the method of the invention, instead of hydroxide, allows the electrodeposition of the sol-gel onto a substrate.
Accordingly, preferably the sol is a ceramic oxide sol, such as titanium oxide.
Zirconia ceramics may also be used as the sols. Gheorghies C. et al (Analele Stiintifice Ale Universitatii, Tomul XLV-XLVI, s. Fizica Starii Condensate (1999-2000) pages 268-275) discloses cathodic depositions of zirconia sol-gels. The electrical current was used to produce hydroxide ions which caused the deposition of the film. Preferably the acidified sol is ZrO(NO3).
Alternatively, the sol-gel layer can comprise a matrix of a hydrogel with polymers, such as polyvinyl, polyacrylamide or polypyrrole-based polymers, or sugar-derivatised polypyrroles, and their derivatives.
Preferably the sol is a silane. Preferably it has a general formula:
or (R9O)3—Si—CH2—CH2—Si—(OR9)3
where:                R1=straight chain, branched chain, cyclic, non-cyclic, saturated or non-saturated, substituted or non-substituted alkyl; substituted or non-substituted aryl; —NR5; and —COR6; the alkyl preferably contains 1, 2, 3, 4, 5 or 6 carbon atoms;        R2, R3 and R4 are independently selected from; straight chain, branched chain, cyclic, non-cyclic, saturated or non-saturated alkyl; —COR6; —O-alkyl; and —O—COR6; —R7R8; R7N(R6)2 and R7NHR6R8; preferably containing 1, 2, 3, 4, 5, or 6 carbon atoms;        R5=branched or non-branched cyclic or non-cyclic, saturated or non-saturated alkyl; or a benzyl group, preferably containing 1, 2, 3, 4, 5, or 6 carbon atoms;        R6=C1 to C3 alkyl;        R7=C1 to C6 alkyl, especially C1, C2 or C3 alkyl;        R8=Epoxy, —NH2 or —SH; especially
                R9=Straight or branched C1 to C6 alkyl, especially C1, C2 or C3 alkyl.
Preferably R1 is methyl, ethyl, propyl, —NCHCH2CH3, —NC(CH3)CH2CH3,

Preferably R2, R3 and R4 are each independently methyl, ethyl, propyl, —O-methyl, —O-ethyl, —O-propyl, —CHCH2, benzyl, —OCOCH3, or aminoalkyl. Preferably, the aminoalkyl is aminopropyl, especially 3 aminopropyl.
Preferably the silanes are selected from:
Tetramethoxysilane (TMOS), tetraethoxysilane (TEOS), tetrapropoxysilane, methyltrimethoxysilane, methyltriethoxysilane, methyltris(methylethylketoxime)silane (MOS), methyltris(acetoxime)silane, dimethyldi(methylethylketoxime)silane, trimethyl(methylethylketoxime)silane, vinyltris(methylethylketoxime)silane (VOS), methyl tris(methylisobutylketoxime)silane, methylvinyldi(methylethylketoxime)silane, methylvinyldi(cyclohexanoneoxime)silane, vinyltris(methylisobutylketoxime)silane, phenyltris(methylethylketoxime)silane (POS), methyltriacetoxysilane and tetraacetoxysilane.
Preferably:
R1=—CH3, R2=—OCH3, R3=—OCH3 R4=

R1=—CH3, R2=—OCH3, R3=—OCH3, R4=—(CH2)3NH2 (APTMOS);
R1=—CH2CH3, R2=—OCH2CH3, R3=—O—CH2CH3, R4=—(CH2)3NH2 (APTEOS);
R1=—CH3, R2=—OCH3 R3=—OCH3, R4=—(CH2)3SH (MPTMOS);
R9=ethyl (Bis-TEOS); or R9=methyl (Bis-TMOS)
Most preferably the silane is tetramethoxysilane (TMOS).
Such silanes are known in the art and available commercially. Two or more silanes may be used.
Preferably a silane coupling agent is added. Such coupling agents contain a reactive group such as an amine, sulphydryl, oxy, acrylate, vinyl or chloro group. This may be added to the acidified sol suspension or to the neutralised sol suspension. These have been found to improve the stability of the film. Preferred coupling agents include:                3-Aminopropyl dimethyl methoxysilane (MMS)        Aminopropyltriethoxysilane        Aminopropyltrimethoxysilane        Aminopropylmethyldiethoxysilane        Aminopropylmethyldimethoxysilane        Aminoethylaminopropyltrimethoxysilane        Aminoethylaminopropyltriethoxysilane        Aminoethylaminopropylmethyldimethoxysilane        Diethylenetriaminopropyltrimethoxysilane        Diethylenetriaminopropyltriethoxysilane        Diethylenetriaminopropylmethyldimethoxysilane        Diethylenetriaminopropylmethyldiethoxysilane        Cyclohexylaminopropyltrimethoxysilane        Hexanediaminomethyldiethoxysilane        Anilinomethyltrimethoxysilane        Anilinomethyltriethoxysilane        Diethylaminomethyltriethoxysilane        (Diethylaminomethyl)methyldiethoxysilane        Methylaminopropyltrimethoxysilane        Bis(triethoxysilylpropyl)tetrasulfide        Bis(triethoxysilylpropyl)disulfide        Mercaptopropyltrimethoxysilane        Mercaptopropyltriethoxysilane        Mercaptopropylmethyldimethoxysilane        3-thiocyanopropyltriethoxysilane        Glycidoxypropyltrimethoxysilane        Glycidoxypropyltriethoxysilane        Glycidoxypropylmethyldiethoxysilane        Glycidoxypropylmethyldimethoxysilane        Methacryloxypropyltrimethoxysilane        Methacryloxypropyltriethoxysilane        Methacryloxypropylmethyldimethoxysilane        Chloropropyltrimethoxysilane        Chloropropyltriethoxysilane        Chloromethyltriethoxysilane        Chloromethyltrimethoxysilane        Dichloromethyltriethoxysilane        Vinyltrimethoxysilane        Vinyltriethoxysilane        Vinyltris(2-methoxyethoxy)silane        Trimethoxysilyl aminopropyl aminoethyl triacetic acid -(MeO)3SiCH2CH2CH2N(CH2COOH)CH2CH2N(CH2COOH)2
MMS has been found to be especially useful in adenosine sensors and stops the matrix becoming too dense. It is therefore preferably used for such sensors.
An electrical potential of about −900 to about −1500 mV, especially about −900 to about −1200 mV, is preferably used. The electrical potential may be applied for about 10 to about 120 seconds, especially about 20 to about 60 seconds, most preferably about 20 to about 40 seconds, depending on the thickness of sol-gel required.
Preferably the coupling agent is 3-aminopropyltrimethoxysilane (APTMOS).
Glycidylpropyltrimethoxysilane may also preferably be added to the neutralised sol-suspension.
The silane coupling agents preferably comprise one or more, especially two or more reactive groups, such as amine groups that cross-link the silane moieties.
Initial results show that the presence of mercaptan-containing silanes, such as MPTMOS, improve the sensitivity of biosensors produced by this technique, for example by lowering the sensitivity of the biosensor to contaminants in an assay mixture. They also improve the stability of biosensors in storage.
Initial results also show that the addition of bifunctional silanes, such as Bis-TEOS or Bis-TMOS, improve the stability and mechanical strength of biosensors.
Accordingly, the sol-gel is preferably obtained from a mixture comprising a mercapto-containing silane and/or a bifunctional silane.
The inventors have found that the sol-gel may be stabilised by the addition of one or more stabilisers such as a polyhydroxyalcohol. Examples include glycerol, polyethylene glycol (PEG) or polyvinyl alcohol; polysaccharides, such as dextran or chitosan; polyalkylene imine; or sugars, such as mannitol, gluconate, gluconolactone, trehalose, lactitol or sucrose, and benzylalkonium chloride (BAC).
Polyethyleneimine (PEI) may also be added.
Optionally, the biosensor according to the invention may also comprise a further layer of matrix, optionally without any biological material, which may be on top of the matrix layer containing the biological material(s).
Two or more biological materials, such as enzymes, may be used in a cascade; such enzymes may be mixed together and applied at the same time. Alternatively they may be applied as separate layers. A first layer is applied by switching on the electrical potential to form a first layer of matrix. The electrically conductive substrate is placed in a second neutralised matrix suspension containing a second enzyme, and a second matrix layer is applied by switching on an electrical potential again. The matrix used in the second suspension may be the same as or different to the first suspension.
Usually, stability, selectivity and sensitivity are the important factors in the development of a biosensor. As well as the pH stability shown in the inset in FIG. 3, the RP modified biosensor also shows a stable response to a substance in successive detection experiments. FIG. 6 shows an FIA response of a typical hypoxanthine biosensor to addition of 10 μM hypoxanthine solution. Over a one hour test, the biosensor exhibits a stable, reproducible response to the hypoxanthine.
As is known in the art, the activity of biological material(s) trapped within matrix layers of biosensors does not tend to fall. This is attributed to the rigidity and porosity of the sol-gel type matrices. Correspondingly, the activity of biological material(s) in the outer matrix layer of an RP modified biosensor is not affected by the RP layer during long term storage.
Plasma was employed to test the selectivity of the biosensor. FIG. 7 shows the amperometric response of the RP modified biosensor to ATP (40 μM), plasma, and mixtures of plasma and ATP (40 μM). It can be seen that the plasma gives hardly any interfering signal on the RP modified biosensor, indicating that the interferences are eliminated at a low operational potential. The RP modified biosensor of the invention therefore also exhibits good selectivity.
In normal biosensors, plasma interferes in the analysis of the substance which is being detected, as the components of plasma, such as urates and ascorbates generate large responses in the biosensors. Using the biosensor of the invention, these responses are greatly minimised due to the enhanced selectivity. Similarly, extremely reactive catechols at concentrations of 1 mM only generate responses which are between background and normal levels, and significantly less than those obtained from target substances.
Furthermore, the electrochemical behaviour of RP is substantially unaffected by the pH of the supporting electrolyte. This is shown in the cyclic voltammogram of FIG. 3 where the positions of the peaks I and II do not vary significantly with the change in pH. However, the stability and activity of the PB modified biosensors are seriously affected at alkaline pH, which restricts their use to only neutral and acidic solutions.
FIG. 8 depicts chrono-amperometric curves of a hypoxanthine biosensor obtained in a phosphate buffer with different concentrations of hypoxanthine, while the inset shows an enlargement of the selection. A sensitivity of 334.17 nA μM−1 cm−2 is obtained from the curves for the hypoxanthine biosensor.
FIG. 9 shows a graph of current Vs. ATP concentration in the calibration of a typical ATP biosensor, while the inset shows the FIA response of a typical biosensor to ATP (40 μM) at an operating potential: 0 mV vs. Ag/AgCl. A clear, strong response to ATP is observed.
A further aspect of the invention provides a process for producing a biosensor according to the invention, comprising providing an electrically conductive substrate, the substrate preferably comprising platinum or a platinum alloy, gold or a gold alloy, or carbon, and depositing thereon;                (i) a first layer formed on the substrate, the first layer comprising Ruthenium Purple;        (ii) a second layer formed on the first layer, the second layer comprising polyaniline or a derivative thereof comprising one or more non-polar substituents; and        (iii) a third layer formed on the second layer, the third layer comprising one or more enzymes trapped in a matrix or adsorbed onto the second layer.
Preferably, the process also involves heating and electrochemical cycling steps. More than about 20 cycles are preferably carried out, more preferably greater than about 30, and most preferably about 40 cycles. The cycling is preferably followed by heating for a period of time, e.g. overnight, preferably at greater than about 50° C., more preferably greater than about 60° C., still more preferably greater than about 70° C., and most preferably up to about 80° C.
Preferably the second layer is at least partially oxidised, for example, by cycling to around +1.3V, for example, for 4 to 7 cycles.
The substrate, polyaniline or its derivatives, matrix and biological materials may be as defined above for the first aspect of the invention.
The process of the invention is extremely controllable, with the layers obtained from the electrodeposition process being substantially the same each time. FIG. 1 shows a typical cyclic voltammogram monitoring the growth of RP on a gold electrode.
Additionally, the use of RP enables the use of particularly small biosensors. More specifically, biosensors which are approximately 50 μM in diameter and about 0.1 to about 2 mm in length are possible, preferably less than 25 μm in diameter and about 300 μm to about 2 mm long.
The operational stability of the biosensors of the invention is in the region of several hours, which is sufficient for the required detection experiments to be carried out.
According to a further aspect of the invention, the biosensor may be used as part of a foetal monitor.
As mentioned above, the ability to monitor hypoxanthine levels provides an indicator of a baby's levels of oxygen during birth and whether a caesarean section is necessary. The biosensor of the invention can be used to test drops of blood taken from the baby's scalp while it is still in the womb. The enzyme in the matrix layer is chosen in this case to be one which metabolises hypoxanthine, such as xanthine oxidase, and thus indicate how much is in the baby's blood.
The biosensor of the present invention requires less fine-tuning than existing tests for hypoxanthine, such as the blood pH test which needs to identify a pH shift of as little as 0.05, and would therefore be more reliable than the existing tests.
Use of the biosensor to detect chemicals such as purines such as hypoxanthine, ATP, adenosine and/or inosine, is also included within the scope of the invention. A method of detecting the amount of a chemical such as purine or a derivative thereof within a tissue or a bodily fluid, comprising exposing a biosensor according to the invention to a sample of the tissue in vivo or in vitro, and detecting an electrical current produced by the biosensor is also provided. Preferably, the tissue is blood, brain, rough or smooth muscle or cardiac tissue. The fluid may be saliva or urine.
A further aspect of the invention provides a kit comprising a biosensor according to the invention for detecting the presence and/or concentration of a chemical such as a purine or a derivative thereof. The kit may comprise means for recording a current from the biosensor in comparison with a reference electrode and may also comprise means for converting the current into an indication of the presence and/or concentration of the chemical to be analysed, such as purine or a derivative thereof. The kit may be used to detect one or more chemicals, e.g. purines, such as hypoxanthine, adenosine, ATP and/or inosine.
Alternatively, the biosensor may be incorporated into a larger biosensor for home use to enable an analyte, such as purine or a derivative thereof, to be monitored, for example in the saliva, blood or urine of a patient.


A tool for replacing a first database node of a database management system by a second database node. The tool receives an indication that the first database node received a data access request for accessing a database shared between the first database node and the second database node. The tool duplicates the data access request at the first database node. Responsive to a determination that the duplicated data access request includes a data changing statement and a previously executed statement, the tool modifies the duplicated data access request to prevent execution of the data changing statement and the previously executed statement at the second database node. The tool executes the modified duplicated data access request at the second database node. The tool replaces, based on a replacement condition being met, the first database node with the second database node in the database management system.

The present invention relates generally to digital computer systems, and more particularly to a database management system.
A database management system (DBMS) is a software package designed to define, manipulate, retrieve and manage data in a database. A DBMS generally manipulates the data itself, the data format, field names, record structure and file structure. It also defines rules to validate and manipulate this data.
Database management systems are set up on specific data handling concepts, as the practice of administrating a database evolves. The earliest databases only handled individual single pieces of specially formatted data. Presently, more evolved systems can handle different kinds of less formatted data and tie them together in more elaborate ways.

International Business Machines Corporation


The invention is a novel soybean cultivar designated LRR969242 with high yield potential and tolerance to Roundup herbicide. The invention relates to seeds of the cultivar LRR969242, plants of the cultivar LRR969242, and to methods for producing a soybean plant produced by crossing the soybean LRR969242 by itself or another soybean genotype.

The present invention relates to a new and distinctive soybean cultivar, designated LRR969242.
There are numerous steps in the development of any novel, desirable plant germplasm. Plant breeding begins with the analysis and definition of problems and weaknesses of the current germplasm, the establishment of program goals, and the definition of specific breeding objectives. The next step is selection of germplasm that possess the traits to meet the program goals. The goal is to combine in a single variety an improved combination of desirable traits from the parental germplasm. These important traits may include higher seed yield, resistance to diseases and insects, better stems and roots, tolerance to drought and heat, and better agronomic quality.
Choice of breeding or selection methods depends on the mode of plant reproduction, the heritability of the trait(s) being improved, and the type of cultivar used commercially (e.g., F1 hybrid cultivar, pureline cultivar, etc.). For highly heritable traits, a choice of superior individual plants evaluated at a single location will be effective, whereas for traits with low heritability, selection should be based on mean values obtained from replicated evaluations of families of related plants. Popular selection methods commonly include pedigree selection, modified pedigree selection, mass selection, and recurrent selection.
The complexity of inheritance influences choice of the breeding method. Backcross breeding is used to transfer one or a few favorable genes for a highly heritable trait into a desirable cultivar. This approach has been used extensively for breeding disease-resistant cultivars. Various recurrent selection techniques are used to improve quantitatively inherited traits controlled by numerous genes. The use of recurrent selection in self-pollinating crops depends on the ease of pollination, the frequency of successful hybrids from each pollination, and the number of hybrid offspring from each successful cross.
Each breeding program should include a periodic, objective evaluation of the efficiency of the breeding procedure. Evaluation criteria vary depending on the goal and objectives, but should include gain from selection per year based on comparisons to an appropriate standard, overall value of the advanced breeding lines, and number of successful cultivars produced per unit of input (e.g., per year, per dollar expended, etc.).
Promising advanced breeding lines are thoroughly tested and compared to appropriate standards in environments representative of the commercial target area(s) for three or more years. The best lines are candidates for new commercial cultivars; those still deficient in a few traits may be used as parents to produce new populations for further selection.
These processes, which lead to the final step of marketing and distribution, usually take from eight to 12 years from the time the first cross is made. Therefore, development of new cultivars is a time-consuming process that requires precise forward planning, efficient use of resources, and a minimum of changes in direction.
A most difficult task is the identification of individuals that are genetically superior, because for most traits the true genotypic value is masked by other confounding plant traits or environmental factors. One method of identifying a superior plant is to observe its performance relative to other experimental plants and to a widely grown standard cultivar. If a single observation is inconclusive, replicated observations provide a better estimate of its genetic worth.
The goal of a plant breeding is to develop new, unique and superior soybean cultivars and hybrids. The breeder initially selects and crosses two or more parental lines, followed by repeated selfing and selection, producing many new genetic combinations. The breeder can theoretically generate billions of different genetic combinations via crossing, selfing and mutations. The breeder has no direct control at the cellular level. Therefore, two breeders will never develop the same line, or even very similar lines, having the same soybean traits.
Each year, the plant breeder selects the germplasm to advance to the next generation. This germplasm is grown under unique and different geographical, climate and soil conditions, and further selection are then made, during and at the end of the growing season. The cultivars which are developed are unpredictable. This unpredictability is because the breeder""s selection occurs in unique environments, with no control at the DNA level (using conventional breeding procedures), and with millions of different possible genetic combinations being generated. A breeder of ordinary skill in the art cannot predict the final resulting lines he develops, except possibly in a very gross and general fashion. The same breeder cannot produce the same cultivar twice by using the exact same original parents and the same selection techniques. This unpredictability results in the expenditure of large amounts of research monies to develop superior new soybean cultivars.
The development of new soybean cultivars requires the development and selection of soybean varieties, the crossing of these varieties and selection of superior hybrid crosses. The hybrid seed is produced by manual crosses between selected male-fertile parents or by using male sterility systems. These hybrids are selected for certain single gene traits such as pod color, flower color, pubescence color or herbicide resistance which indicate that the seed is truly a hybrid. Additional data on parental lines as well as the phenotype of the hybrid influence to breeder""s decision whether to continue with the specific hybrid cross.
Pedigree breeding and recurrent selection breeding methods are used to develop cultivars from breeding populations. Breeding programs combine desirable traits from two or more cultivars or various broad-based sources into breeding pools from which cultivars are developed by selfing and selection of desired phenotypes. The new cultivars are evaluated to determine which have commercial potential.
Pedigree breeding is used commonly for the improvement of self-pollinating crops. Two parents which possess favorable, complementary traits are crossed to produce an F1. An F2 population is produced by selfing one or several F""s. Selection of the best individuals may begin in the F2 population; then, beginning in the F3, the best individuals in the families are selected. Replicated testing of families can begin the F4 generation to improve the effectiveness of selection for traits with low heritability. At an advanced stage of inbreeding (i.e., F6 and F7), the best lines or mixtures of phenotypically similar lines are tested for potential release as new cultivars.
Mass and recurrent selections can be used to improve populations of either self- or cross-pollinating crops. A genetically variable population of heterozygous individuals is either identified or created by intercrossing several different parents. The best plants are selected based on individual superiority, outstanding progeny, or excellent combining ability. The selected plants are intercrossed to produce a new population in which further cycles of selection are continued.
Backcross breeding has been used to transfer genes for a simply inherited, highly heritable trait into a desirable homozygous cultivar or inbred line which is the recurrent parent. The source of the trait to be transferred is called the donor parent. The resulting plant is expected to have the attributes of the recurrent parent (e.g., cultivar) and the desirable trait transferred from the donor parent. After the initial cross, individuals possessing the phenotype of the donor parent are selected and repeatedly crossed (backcrossed) to the recurrent parent. The resulting plant is expected to have the attributes of the recurrent parent (e.g., cultivar) and the desirable trait transferred from the donor parent.
The single-seed descent procedure in the strict sense refers to planting a segregating population, harvesting a sample of one seed per plant, and using the one-seed sample to plant the next generation. When the population has been advanced from the F2 to the desired level of inbreeding, the plants from which lines are derived will each trace to different F2 individuals. The number of plants in a population declines each generation due to failure of some seeds to germinate or some plants to produce at least one seed. As a result, not all of the F2 plants originally sampled in the population will be represented by a progeny when generation advance is completed.
In a multiple-seed procedure, soybean breeders commonly harvest one or more pods from each plant in a population and thresh them together to form a bulk. Part of the bulk is used to plant the next generation and part is put in reserve. The procedure has been referred to as modified single-seed descent or the pod-bulk technique.
The multiple-seed procedure has been used to save labor at harvest. It is considerably faster to thresh pods with a machine than to remove one seed from each by hand for the single-seed procedure. The multiple-seed procedure also makes it possible to plant the same number of seeds of a population each generation of inbreeding. Enough seeds are harvested to make up for those plants that did not germinate or produce seed.
Descriptions of other breeding methods that are commonly used for different traits and crops can be found in several reference books (e.g., Allard, 1960; Simmonds, 1979; Sneep et al., 1979; Fehr, 1987).
Proper testing should detect any major faults and establish the level of superiority or improvement over current cultivars. In addition to showing superior performance, there must be a demand for a new cultivar that is compatible with industry standards or which creates a new market. The introduction of a new cultivar will incur additional costs to the seed producer, the grower, processor and consumer, for special advertising and marketing, altered seed and commercial production practices, and new product utilization. The testing proceeding release of a new cultivar should take into consideration research and development costs as well as technical superiority of the final cultivar. For seed-propagated cultivars, it must be feasible to produce seed easily and economically.
Soybean, Glycine max (L), is an important and valuable field crop. Thus, a continuing goal of plant breeders is to develop stable, high yielding soybean cultivars that are agronomically sound. The reasons for this goal are obviously to maximize the amount of grain produced on the land used and to supply food for both animals and humans. To accomplish this goal, the soybean breeder must select and develop soybean plants that have the traits that result in superior cultivars.
The invention is a novel soybean cultivar designated LRR969242 with high yield potential and tolerance to Roundup herbicide. The invention relates to seeds of the cultivar LRR969242, plants of the cultivar LRR969242, and to methods for producing a soybean plant produced by crossing the soybean LRR969242 by itself or another soybean genotype.
The invention is also directed to soybean cultivar LRR969242 further comprising one or more specific, single gene traits, for example transgenes, and which has essentially all of the morphological and physiological characteristics of cultivar LRR969242, in addition to the one or more specific, single gene traits. The invention further relates to seeds of cultivar LRR969242 further comprising one or more specific, single gene traits, plants of cultivar LRR969242 further comprising one or more specific, single gene traits, and to methods for producing a soybean plant by crossing of a soybean plant of cultivar LRR969242 further comprising one or more specific, single gene traits by itself or another soybean genotype.
In the description and tables that follow, a number of terms are used. In order to provide a clear and consistent understanding of the specification and claims, including the scope to be given such terms, the following definitions are provided:
Seed Yield (Bushels/Acre). The yield in bushels/acre is the actual yield of the grain at harvest.
Maturity Date. Plants are considered mature when 95% of the pods have reached their mature color.
Lodging Resistance. Lodging is rated on a scale of 1 to 9. Where one is completely upright and 9 is completely prostrate.
Plant Height. Plant height is taken from the top of soil to top of node of the plant and is measured in centimeters.
Iron-Deficiency Chlorosis. Plants are scored 1 to 9 based on visual observations. A score of 1 means no stunting of the plants or yellowing of the leaves and a score of 9 indicates the plants are dead or dying caused by iron-deficiency chlorosis.
Shatter. The amount of pod dehiscence prior to harvest. Pod dehiscence involves seeds falling from the pods to the soil. This is a visual score from 1 to 9 comparing all genotypes within a given test. A score of 1 means pods have not opened and no seeds have fallen out. A score of 9 indicates 100% of the pods are opened.


Homeotropically oriented layers and films of liquid crystals especially on polymeric substrates. The layers and films, which also can be multi-layered films comprising a homeotropically aligned anisotropic film, are preparable by application of an orientation layer to the substrate. The orientation layer can be either an organic surfactant which is fixed in a matrix of a polymeric material or an inorganic thin layer such as aluminum or Al2O3.

This work aims to find a suitable homeotropic alignment method for orthogonal liquid crystals, especially for nematic and smectic A liquid crystal mixtures on plastic substrates.
A preferred application is the orientation of reactive liquid crystal mixtures for the preparation of anisotropic polymeric films.
The reactive liquids crystal are dissolved in a suitable solvent such as toluene and coated as a thin film onto a flexible plastic substrate. When the solvent evaporates and the reactive liquid crystal remains, the liquid crystal has to align instantly, so that high speed roll to roll coating of the plastic substrate is possible. This is achieved by the method described hereafter. The reactive liquid crystal is subsequently polymerised using UV light, which is acting upon a photoinitiator mixed into the reactive liquid crystal, or alternatively by heat, which in turn is acting upon a thermal initiator mixed into the reactive liquid crystal. A polymer film having oriented constituent molecules i.e. monomer molecules is thus produced. It is well known that photopolymerisation of aligned reactive liquid crystals gives rise to polymer films having molecules aligned in the same way as the monomer reactive liquid crystal, thus freezing or stabilizing the director orientation of the liquid crystal and preserving it""s anisotropic properties. Such films are characterized by and useful for their optical properties. They are uniaxial, positive birefringent films.
Homeotropic alignment of a liquid crystal occurs when the long axes of the molecules of the liquid crystal phase are, on average, essentially normal to the substrate upon which they form a thin film. Very few materials spontaneously align in this way and thus some kind of aligning agent is required to produce this desired alignment. Orthogonal liquid crystal phases like nematic liquid crystal phases and orthogonal smectic phases (e.g. smectic A and smectic B) can be aligned in this way. An Overview about orientation technologies for liquid crystals is given e.g. in J. Cognard, Molecular Crystals Liquid Crystals 78 Supplement 1 (1981) pp 1-77 and in JA. Castellano MCLC, 94 (1983) pp 33-41.
Conventional aligning agents for liquid crystals are designed to be effective on glass substrates, some examples of such conventional organic aligning agents are lecithin, trichloro- and trimethoxypropyl silane, hexadecyl trimethyl ammonium halides and alkyl carboxylato monochromium salts. In each case a very small amount (of typically less than 1%) of the active component is dissolved in a suitable volatile solvent and spread onto the substrate e.g. by spin coating or other well known methods, when the solvent has been evaporated, a thin film of the organic aligning agent remains on the substrate. These materials are characterized by having a polar end group which is assumed to be attracted to the polar glass surface and a long alkyl chain which orientates normal to the glass surface. In some cases a chemical reaction is essential i.e. in the case of chlorosilanes which react with hydroxyl groups on the glass surface. Spontaneous homeotropic alignment of liquid crystals occurs on such surfaces.
Inorganic aligning agents can also be used. For example it is well known that evaporation at normal incidence of SiO2 or MgF2 onto a substrate of glass can be used to produce homeotropic alignment of liquid crystals.
This patent application, however, relates to the alignment of orthogonal liquid crystal phases especially of nematic and of smectic A liquid crystals on a plastic substrate. The plastic substrates are preferably flexible substrates and may be plastic films or also anisotropic liquid crystal polymer films. It was found that the conventional aligning agents are not effective for the alignment of liquid crystals on such plastic substrates. Typically and almost generally either no alignment or only very poor alignment is exhibited. It is suspected that the surface of the plastic substrate has much less affinity for the polar end groups of the aligning agents.
In contrast to the conventionally used organic aligning agents, inorganic aligning agents such as typically used on glass surfaces e.g. SiO2 have been found to provide good homeotropic alignment when sputter coated or evaporated onto plastic substrates. It was found, however, that TiO2-coated substrates do not align so well and are more difficult to wet with the usual coating solvents. One of the problems with these types of coatings which is common to SiO2, TiO2 and others, is that they tend to have a significant surface roughness which prevents the polymer film being released from the aligning layer after polymerisation. This could be attributed to the well known fact that adhesion is very much enhanced by using microscopically rough surfaces compared to smoother surfaces. As these coatings are usually deposited onto a high temperature stable substrate such as polyethylene terephtalate (PET) which, due to the stretching processes used in its manufacture, is birefringent the aligned polymer film must be removed from the substrate before the optical properties of the coated polymer itself are accessible.
This invention relates to a technique which can be used to quickly align orthogonal liquid crystals especially nematic and smectic A liquid crystals on plastic substrates and which is applicable to roll to roll coating of plastic substrates. It was found that certain organic and inorganic surfactants can be used to provide homeotropic alignment on plastic substrates. This will be explained in the following.
One type of such organic surfactants is the family of perfluoroalkyl sulphonates represented by the surfactant FC 431 (tradename of commercial product obtainable from the 3M company, USA) which is a liquid dissolved in a solvent. When added to the reactive liquid crystal (20% solids), in toluene at a concentration of from 0.1 to 0.5% and preferred at a concentration of from 0.7 to 0.4% it promotes homeotropic alignment on plastic surfaces such as triacetyl cellulose (TAC), (available from the Lonza company in Switzerland) or polyethylene terephthalate (PET), (available from the ICI company in England under the tradename of Melinex 401). To improve the alignment, corona discharge (CD) treatment of the substrate has been found to be helpful. This possibly could be attributed to the CD treatment increasing the polarity of the surface by the formation of hydroxyl and carbonyl groups. However, the alignment process even with a CD treatment is rather slow, taking some seconds after the solvent has evaporated, which has a detrimental effect on the speed at which any coating process can proceed and consequently is not acceptable. The aligning agent has to migrate from the bulk to the surface which appears to have little attraction for it. Thus the alignment process is both weak and slow.
A further topic of the present invention is to induce this type of surfactant to align faster whilst also making it into a form that is suitable for roll to roll coating applications. As the surfactant is a liquid and thus it is practically not possible to coat it onto the surface of the substrate if the substrate is in the form of a roll, unless two or more coating stations are available. However, surprisingly if the FC431 is embedded in a UV curable polymer C3 film which is coated onto the substrate and subsequently UV cured, the resultant polymer film with the surfactant retains its alignment capabilities and the alignment is now much faster and thus allowing a highly desirable greater coating speed.
We found that surfactants can be fixed on the surface e.g. by making e.g. a mixture of 5% surfactant in an acrylate monomer mixture such as a mixture of hexanediol diacrylate and ethylhexylacrylate and 4% Irgacure 907 and coating this onto the PET (or TAC) film and then photocuring the mixture. This provides a faster alignment.
The active polymerizable component can be a monomer or a mixture of monomers or a mixture of monomers and oligomers. They can be anisotropic but preferably are isotropic compounds.
The preferred concentration range for the surfactant in the monomer or monomer mixture or monomer oligomer mixture has been found to be from 2 to 8% surfactant. Especially preferred are concentrations of from 4 to 6%.
Suitable UV curable polymers which are useful to embed the aligning agent can be made from mixtures of known available mono-, di- and tri-acrylates such as 2-methyl hexylacrylate and hexane diol diacrylate. It is further often helpful to also incorporate mono- and diacrylated oligomers which can be based on all the common materials such as urethanes or esters. A wide range of such compounds is available from UCB, Belgium and other suppliers of UV curable resins. These other reactive compounds are used to improve other properties such as better durability, flexibility, hardness and different surface energies to aid wetting.
In a preferred embodiment of the invention the reactive mesogenic compounds used in the mixture of the polymerizable mesogenic material are compounds of formula I
P-(Sp-X)n-MG-Rxe2x80x83xe2x80x83I
wherein
P is a polymerizable group,
Sp is a spacer group having 1 to 20 C atoms,
X is a group selected from xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94COxe2x80x94, xe2x80x94COOxe2x80x94, xe2x80x94OCOxe2x80x94, xe2x80x94OCOxe2x80x94Oxe2x80x94 or a single bond,
n is 0 or 1,
MG is a mesogenic or mesogenity supporting group, preferably selected according to formula II
-(A1-Z1)m-A2-Z2-A3-xe2x80x83xe2x80x83II
wherein
A1, A2
and A3 are independently from each other 1,4-phenylene in which, in addition, one or more CH groups may be replaced by N, 1,4-cyclohexylene in which, in addition, one or two non-adjacent CH2 groups may be replaced by O and/or S, 1,4-cyclohexenylene or naphthalene-2,6-diyl, it being possible for all these groups to be unsubstituted, mono- or poly-substituted with halogen, cyano or nitro groups or alkyl, alkoxy or acyl groups having 1 to 7 C atoms wherein one or more H atoms may be substituted by F or Cl,
Z1 and Z2 are each independently xe2x80x94COOxe2x80x94, xe2x80x94OCOxe2x80x94, xe2x80x94CH2CH2xe2x80x94, xe2x80x94OCH2xe2x80x94, xe2x80x94CH2Oxe2x80x94, xe2x80x94CHxe2x95x90CHxe2x80x94, xe2x80x94Cxe2x89xa1Cxe2x80x94, xe2x80x94CHxe2x95x90CHxe2x80x94COOxe2x80x94, xe2x80x94OCOxe2x80x94CHxe2x95x90CHxe2x80x94 or a single bond and
m is 0, 1 or 2,
and
R is an alkyl radical with up to 25 C atoms which may be unsubstituted, mono- or polysubstituted by halogen or CN, it being also possible for one or more non-adjacent CH2 groups to be replaced, in each case independently from one another, by xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94NHxe2x80x94, xe2x80x94N(CH3)xe2x80x94, xe2x80x94COxe2x80x94, xe2x80x94COOxe2x80x94 xe2x80x94OCOxe2x80x94, xe2x80x94OCOxe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94COxe2x80x94, xe2x80x94COxe2x80x94Sxe2x80x94 or xe2x80x94Cxe2x89xa1Cxe2x80x94 in such a manner that oxygen atoms are not linked directly to one another, or alternatively R is halogen, cyano or has independently one of the meanings given for P-(Sp-X)n-.
Particularly preferred are polymerizable mixtures comprising at least two reactive mesogenic compounds at least one of which is a compound of formula I.
In another preferred embodiment of the invention the reactive mesogenic compounds are selected according to formula I, wherein R has one of the meanings of P-(Sp-X)nxe2x80x94 given above.
Mesogenic compounds containing two or three rings are preferred.
Halogen is preferably F or Cl, most preferred it is F.
Of the compounds of formula I especially preferred are those in which R is F, Cl, cyano, alkyl or alkoxy or has the meaning given for P-(Sp-X)n-, and MG is of formula II wherein Z1 and Z2 are xe2x80x94COOxe2x80x94, xe2x80x94OCOxe2x80x94, xe2x80x94CH2xe2x80x94CH2xe2x80x94, xe2x80x94CHxe2x95x90CHxe2x80x94COOxe2x80x94, xe2x80x94OCOxe2x80x94CHxe2x95x90CHxe2x80x94 or a single bond.
A smaller group of preferred mesogenic groups of formula II is listed below. For reasons of simplicity, Phe in these groups is 1,4-phenylene, PheL is a 1,4-phenylene group which is substituted by at least one group L, with L being F, Cl, CN or an optionally fluorinated alkyl, alkoxy or acyl group with 1 to 4 C atoms preferably L is F or CH3, and Cyc is 1,4-cyclo-hexylene.
In these preferred groups Z1 and Z2 have the meaning given in formula II described above. Preferably Z1 and Z2 are xe2x80x94COOxe2x80x94, xe2x80x94OCOxe2x80x94, xe2x80x94CH2CH2xe2x80x94 or CHxe2x95x90CHxe2x80x94COOxe2x80x94. L is preferably F, Cl, CN or methyl, methoxy, ethyl, ethoxy, oxamethyl, oxaethyl or trifluormethyl.
If R is an alkyl or alkoxy radical, i.e. where the terminal CH2 group is replaced by xe2x80x94Oxe2x80x94, this may be straight-chain or branched. It is preferably straight-chain, has 2, 3, 4, 5, 6, 7 or 8 carbon atoms and accordingly is preferably ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, ethoxy, propoxy, butoxy, pentoxy, hexoxy, heptoxy, or octoxy, furthermore methyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, methoxy, nonoxy, decoxy, undecoxy, dodecoxy, tridecoxy or tetradecoxy.
Oxaalkyl, i.e. where one CH2 group is replaced by xe2x80x94Oxe2x80x94, is preferably straight-chain 2-oxapropyl (xe2x95x90methoxymethyl), 2- (xe2x95x90ethoxymethyl) or 3-oxabutyl (xe2x95x902-methoxyethyl), 2-, 3-, or 4-oxapentyl, 2-, 3-, 4-, or 5-oxahexyl, 2-, 3-, 4-, 5-, or 6-oxaheptyl, 2-, 3-, 4-, 5-, 6- or 7-oxaoctyl, 2-, 3-, 4-, 5-, 6-, 7- or 8-oxanonyl or 2-, 3-, 4-, 5-, 6-, 7-, 8- or 9-oxadecyl.
R optionally can be a chiral group. Compounds with a chrial group R in most cases, however, are not preferred in mixtures with a nematic phase which are the preferred mixtures according to the instant invention. In case of R being a chiral group it is preferably selected of the following formula
wherein
X1 has the meaning given for X,
Q1 is an alkylene or alkyleneoxy group with 1 to 10 C atoms or a single bond,
Q2 is an alkyl or alkoxy group with 1 to 10 C atoms which may be unsubstituted, mono- or polysubstituted by halogen or CN, it being also possible for one or more non-adjacent CH2 groups to be replaced, in each case independently from one another, by xe2x80x94Cxe2x89xa1Cxe2x80x94, xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94NHxe2x80x94, xe2x80x94N(CH3)xe2x80x94, xe2x80x94COxe2x80x94, xe2x80x94COOxe2x80x94, xe2x80x94OCOxe2x80x94, xe2x80x94OCOxe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94COxe2x80x94 or xe2x80x94COxe2x80x94Sxe2x80x94 in such a manner that oxygen atoms are not linked directly to one another, or alternatively has the meaning given for P-Sp-,
Q3 is halogen, a cyano group or an alkyl or alkoxy group with 1 to 4 C atoms different from Q2.
In case chiral groups are used the preferred chiral groups are 2-butyl (xe2x95x901-methylpropyl), 2-methylbutyl, 2-methylpentyl, 3-methylpentyl, 2-ethylhexyl, 2-propylpentyl, 2-octyl, in particular 2-methylbutyl, 2-methylbutoxy, 2-methylpentoxy, 3-methylpentoxy, 2-ethylhexoxy, 1-methylhexoxy, 2-octyloxy, 2-oxa-3-methylbutyl, 3-oxa-4-methylpentyl, 4-methylhexyl, 2-nonyl, 2-decyl, 2-dodecyl, 6-methoxyoctoxy, 6-methyloctoxy, 6-methyloctanoyloxy, 5-methylheptyloxycarbonyl, 2-methylbutyryloxy, 3-methylvaleroyloxy, 4-methylhexanoyloxy, 2-chlorpropionyloxy, 2-chloro-3-methylbutyryloxy, 2-chloro-4-methylvaleryloxy, 2-chloro-3-methylvaleryloxy, 2-methyl-3-oxapentyl, 2-methyl-3-oxahexyl, 1-methoxypropyl-2-oxy, 1-ethoxypropyl-2-oxy, 1-propoxypropyl-2-oxy, 1-butoxypropyl-2-oxy, 2-fluorooctyloxy, 2-fluorodecyloxy, for example.
In addition, mesogenic compounds of the formula I containing an achiral branched group R may occasionally be of importance as comonomers, for example, as they reduce the tendency towards crystallization. Branched groups of this type generally do not contain more than one chain branch. Preferred achiral branched groups are isopropyl, isobutyl (xe2x95x90methylpropyl), isopentyl (xe2x95x903-methylbutyl), isopropoxy, 2-methylpropoxy and 3-methylbutoxy.
It his however also possible to use reactive compounds with chiral groups selected from the following groups:
a cholesteryl group,
an ethylenglycol derivative
wherein R1 is an alkyl radical with 1 to 12 C atoms,
a terpenoid radical like, for example, menthol
or a group based on citronellol.
Preferably R is an achiral group. In orthogonal smectic phases the influence of chiral compounds generally is rather small.
P is preferably selected form CH2xe2x95x90CW-COOxe2x80x94, WCHxe2x95x90CHxe2x80x94Oxe2x80x94,
or CH2xe2x95x90CH-Phenyl-(O)kxe2x80x94 with W being H, CH3 or Cl and k being 0 or 1,
P is particularly preferably a vinyl group, an acrylate group, a methacrylate group, a propenyl ether group or an epoxy group, very particularly preferably an acrylate group.
As for the spacer group Sp all groups can be used that are known for this purpose to those skilled in the art. The spacer group Sp is preferably linked to the polymerizable group P by an ester or ether group or a single bond. The spacer group Sp is preferably a linear or branched alkylene group having 1 to 20 C atoms, in particular 1 to 12 C atoms, in which, in addition, one or more non-adjacent CH2 groups may be replaced by xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94NHxe2x80x94, xe2x80x94N(CH3)xe2x80x94, xe2x80x94COxe2x80x94, xe2x80x94Oxe2x80x94COxe2x80x94, xe2x80x94Sxe2x80x94COxe2x80x94, xe2x80x94Oxe2x80x94COOxe2x80x94, xe2x80x94COxe2x80x94Sxe2x80x94, xe2x80x94COxe2x80x94Cxe2x80x94, xe2x80x94CH(halogen)-, xe2x80x94CH(CN)xe2x80x94, xe2x80x94CHxe2x95x90CHxe2x80x94 or xe2x80x94Cxe2x89xa1Cxe2x80x94.
Typical spacer groups are for example xe2x80x94(CH2)oxe2x80x94, xe2x80x94(CH2CH2O)rxe2x80x94CH2CH2xe2x80x94, xe2x80x94CH2CH2xe2x80x94Sxe2x80x94CH2CH2xe2x80x94 or xe2x80x94CH2CH2xe2x80x94NHxe2x80x94CH2CH2xe2x80x94, with o being an integer from 2 to 12 and r being an integer from 1 to 3.
Preferred spacer groups are ethylene, propylene, butylene, pentylene, hexylene, heptylene, octylene, nonylene, decylene, undecylene, dodecylene, octadecylene, ethyleneoxyethylene, methyleneoxybutylene, ethylene-thioethylene, ethylene-N-methyl-iminoethylene and 1-methyl-alkylene, for example.
In the event that R or Q2 is a group of formula P-Sp-X- or P-Sp- respectively, the spacer groups on each side of the mesogenic core may be identical or different.
In particular, preferred are compounds of formula I wherein n is 1.
In another preferred embodiment, the inventive compensator is obtained by copolymerizing mixtures comprising compounds of formula I wherein n is 0 and compounds of formula I wherein n is 1.
Typical examples representing polymerizable mesogenic compounds of the formula I can be found in WO 93/22397; EP 0,261,712; DE 195,04,224; DE 4,408,171 or DE 4,405,316. The compounds disclosed in these documents, however are to be regarded merely as examples that should not limit the scope of this invention.
Furthermore, typical examples representing polymerizable mesogenic compounds are shown in the following list of compounds, which is, however, to be understood only as illustrative without limiting the scope of the present invention:
In these compounds Nos. 1 to 5 x and y are each independently 1 to 12, A is a 1,4-phenylene or 1,4-cyclohexylene group, R1 is halogen, cyano or an alkyl or alkoxy group with 1 to 12 C atoms and L1 and L2 are each independently H, Halogen, CN, or an alkyl, alkoxy or acyl group with 1 to 7 C atoms.
The reactive mesogenic compounds disclosed in the foregoing and the following can be prepared by methods which are known per se and which are described in the documents cited above and, for example, in standard works of organic chemistry such as, for example, Houben-Weyl, Methoden der organischen Chemie, Thieme-Verlag, Stuttgart.
In a preferred embodiment of the present invention, the compensation film is obtainable from a mixture of a polymerizable mesogenic material comprising the following components
a1) 10 to 99% by weight of at least one mesogen according to formula I and II having one polymerizable functional group,
a2) 0 to 90% by weight of at least one mesogen according to formula I and II having two or more polymerizable functional groups,
b) 0.01 to 5% by weight of an initiator,
c) 0 to 20% by weight of a non-mesogenic compound having two or more polymerizable functional groups,
d) 0 to 1000 ppm of a stabilizer, and
e) 0 to 2% of an organic surfactant.
In a particularly preferred embodiment of the invention the mixture of the polymerizable mesogenic material comprises 15 to 99% by weight of at least one mesogen of component a1) and further comprises at least 5% of at least one mesogen of compound a2), components b) and optionally c) and d) as described above.
In a preferred embodiment the mixture contains at least 0.1% to 1.5% of the organic surfactant, component e) above.
The mixture according to this particularly preferred embodiment preferably comprises one, two or three different mesogens according to formula I and II having one polymerizable functional group.
Most preferably the mixture according to this particularly preferred embodiment comprises two or more, in particular two to six, very particularly two to three different mesogens according to formula I and II having one polymerizable functional group.
The ratio of each of the mesogens according to formula I and II having one polymerizable functional group in the mixture according to this particularly preferred embodiment is preferably 5 to 90, in particular 10 to 80, very preferably 15 to 65% by weight of the total mixture.
In the mixture according to the particularly preferred embodiment described above, preferably each of the different mesogens according to formula I and II is different in at least one of the groups P, Sp, X, Axe2x80x2, A2, A3, Zxe2x80x2, Z2 and R from each other of the mesogens.
The mixture according to this particularly preferred embodiment especially preferably contains from 2 to 10% by weight, very especially preferably from 5 to 10% of at least one of the compounds of component a2).
In another particularly preferred embodiment of the present invention, the mixture of the polymerizable mesogenic material comprises
a1) 15 to 85% by weight of at least one mesogen according to formula I and II having one polymerizable functional group,
a2) 5 to 80% by weight of at least one mesogen according to formula I and II having two or more polymerizable functional groups, and further comprises component b) and optionally component c) and d) as described above.
In one embodiment of the instant invention the organic surfactant is used in a concentration of 0.01 to 1% and especially preferred in 0.1 to 0.5%, in particular 0.1 to 0.3% are used.
In a preferred enbodiment, however, the surfactant is used in a matrix of a polymerizable acrylate monomer or mixture of such monomers and/or oligomers. Then obviously generally no surfactant is required in the mixture of anisotopic reactive monomers which are aligned to form the anisotopic layer or film.
In both embodiments described above sulfonate surfactants are preferred. Especially preferred are fluorinated sulfonate surfactants.
The precursor of the aligning layer contains preferably non mesogenic acrylates and non mesogenic oligomers together with the surfactant and an appropriate initiator, which preferably is a UV initiator. A concentration of from 2 to 10% especially of from 4 to 6% of surfactant in a mixture of monoacrylates and diacrylates and/or oligomers has found to work very well. Especially preferred are mixtures containing at least one monoacrylate, one diacrylate and one oligomer together with the surfactant and the initiator. The concentrations of the monoacrylate in the mixtures are preferably from 10 to 80%, those of the diacrylate monomers from 10 to 50% and those of the oligomers from 10 to 50%. The oligomers are preferably multifunctional poly urethanes or poly esters. The photoinitiator is used in a concentration from 3 to 6%, a typical example of a photoinitiator being Irgacure 907. The mixture is coated from toluene or methyl ethyl ketone. A 10 to 20% solution provides a thin film with a thickness of below 2 xcexcm. The film thickness is not very crucial. However, preferably the film thickness is from 0.5 to 5 xcexcm most preferably from 0.5 to 1.5 xcexcm.
Without further elaboration one skilled in the art can, using the preceding description, utilize the present invention to its fullest extent. The following examples are, therefore, to be construed as merely illustrative and by no means limiting the remainder of the disclosure in any way whatsoever.
In the foregoing and in the following examples, unless otherwise indicated, all temperatures are set forth uncorrected in degrees Celsius and all parts and percentages are by weight. All physical data are given for a temperature of 20xc2x0 C. unless explicitly stated otherwise. The following abbreviations are used to illustrate the liquid crystalline phase behaviour of the compounds:
K=crystalline; N=nematic; S=smectic; Ch=cholesteric; I=isotropic. The numbers between these symbols indicate the phase transition temperatures in degree Celsius.
Other components such as wetting agents and adhesion promoters (available from Cray Valley, USA and Akzo Ltd, Netherlands for example) can also be added to aid coating of the film. Such compounds can also lead to homeotropic alignment. An example for substances which is used as a typical adhesion promoter, which at the same time leads to homeotropic alignment are epoxy acrylate adhesion promoters, e.g. Crodomer UVE-150 from Croda, UK. Typically from 0.01% to 1% of the compound or compounds is/are added, to the mixture of reactive mesogens (the reactive liquid crystal mixture). A preferred concentration for the use of these compounds is from 0.05 to 0.2% and especially about 0.1%.
For UV curable systems photoinitiators such as Irgacure 907, 184, 651 and 369 and Daracur 1173 (all available form Ciba Geigy, Switzerland) have also to be added to create free radicals under the influence of UV light and promote polymerisation of the acrylates. Alternatively thermal photoinitiators such as benzoyl peroxide can also be used if the cure mechanism was to be induced by heat, however this is generally less convenient and consequently not preferred.
Other perfluoroalkyl sulphonate surfactants such as FC430 (also available from the 3M company, USA) also behave in the same way, as FC431 which is given as a typical example of the use of perfluoroalkyl sulphonates only. A further suitable fluorosurfactant is FC 723 also of 3M company. A wide range of similar useful surfactants are available from the 3M company. Also other perfluoro based surfactants can be beneficially used.
Additionally, inorganic aligning agents are well known to be used on glass substrates and when coated onto the substrate surface are also effective aligning agents on plastic substrates, in contrast to conventional organic aligning agents. However, the surface roughness inherent to these inorganic coatings (i.e. using no additional spacers) is a major problem because this together with the high surface energy of the films, which typically is in the range of some tens of dynes e.g. 60 dynes, prevents the film beeing easily released from the surface after UV curing of the reactive liquid crystal. In some cases it is possible to use release agents in the coating solution to aid release of the polymer film. However, these release agents in may cases have an adverse effect on the alignment and in some cases cause a fine liquid deposit to occur in the film. This effect is also called blooming of the film. In order to over come these negative effects preferably no release agents are applied.
It was found that these problems can be overcome by the use of substrates which have been coated with aluminium or sufficiently smooth Al2O3. The substrate can be any suitable plastic such as TAC, polyethylene terepthalate, polyethylene naphthenate, polyamides, and any other plastic; but usually the substrate is PET because it is robust and readily available as a commodity item used extensiveley in the packaging industry. In the case aluminium coated substrates are used the same mixtures of reactive liquid crystals can be used as described above. Here, however, the use of a surfactant is optional only.
Aluminium coated PET is commercially available in many grades of surface finish and with various aluminium thickness and PET thickness. Of necessity, coatings of aluminium have a surface coating of aluminium oxide on them but as the aluminium coating can be obtained in a very smooth form, which is required to provide a desired high reflectivity, the oxide layer is also very smooth, unlike the aluminium oxide coatings obtainable by evaporation methods or by sputtering which all tend to give more porous and rougher surfaces, similar to those which are used e.g. for SiO2 to align liquid crystals on glass substrates. Smoother Al2O3 coatings are also suitable but they are generally less easily available. An exception, however, are the Al2O3 coatings on PET films used in transparent food packaging. Consequently such high reflectivity aluminium coatings and thin transparent Al2O3 coatings, are preferred for the instant invention. Low optical density aluminium coatings, with optical densities from below 0.5 to below 1.0 OD, are not so desirable, because the alignment they induce tends to be not uniform. Thicker, medium and high optical density aluminium coatings are preferred, as these give uniform coatings. Medium density aluminium coatings have an optical density from 1.0 to 2.0 OD, whereas high density aluminium coatings have an optical density of more than 2.0 OD.
The reflective and conductive nature of the aluminium layer has so far precluded its consideration as an alignment layer in liquid crystal displays with glass substrates. For the application described here, these considerations are however less relevant.
Depending on the accuracy and coat weight of the final film a wide range of coating techniques can be used to coat the reactive liquid crystal mixture solution onto the web. A non exhasutive list of examples is: three roll reverse coating, flexo, gravure, micro-gravure, slot die and metered slot die.
The photopolymerisation of the inventive films can be carried out using medium or high pressure mercury lamps which are advantageously filtered using water or dichroic filters to remove light with short wavelengths and reduce the heating effect, as this undesired effect otherwise can cause the reactive liquid crystal mixture to be heated above its clearing point and thus eliminate the liquid crystal phase. Other lamps such as metal halogen lamps and incandescent tubes can also be used but an inert gas atmosphere would typically be required to allow full surface cure in these cases. Typically used are mercury UV lamps with about 80-100 watts/cm irradiation. The emitted light is in the wave length range from 300 to 400 nm.
The UV curing of the aligned polymer film has to proceed at not too high temperatures, as otherwise, especially at longer curing times the optical retardation of the final film will be reduced. Preferred are curing times at from 1 second to 5 minutes and especially from 10 seconds to 2 minutes. The most preferred temperatures during curing are above ambient temperature and about 20 degrees below the clearing point of the reactive liquid crystal mixture. It is generally preferred to cure the mixture as far as possible into the nematic phase (i.e. at as low a temperature as possible). Preferably the curing is performed about at least 65xc2x0 below the clearing point and especially preferred at least 70xc2x0 below the clearing point. Especially preferred are 20xc2x0 C. to 50xc2x0 C. and in particular 20xc2x0 C. to 40xc2x0 C.
The homeotropically aligned polymer films made on aluminised PET or alternatively on polymer/surfactant surfaces can be removed from the PET substrate by pulling them off with a pressure sensitive adhesive coated onto a non-birefringent substrate such as TAC. The alignment and thickness of the films can be quantified by measuring the retardation of the film at some angle away form normal incidence. For a homeotropic film the retardation value is symmetrical about normal incidence. Several methods for measuring retardation are available such as the Berek compensator and polarising microscope (both available form Olympus Microscopes). Qualitatively the film appears black between crossed polarisers.
The reactive liquid crystals can be of any form such that they exhibit nematic or orthogonal smectic phases. However most preferred are the compounds described and listed above.
Besides the acrylates mentioned as examples other reactive groups can also be used such as methacrylates, haloacrylates and epoxies or vinyl ethers. The latter of which will require the use of cationic photoinitiators.
In the case of nematic liquid crystals it is essential to cure the films below the clearing point of the nematic liquid crystal mixture and not to allow the film temperature to reach the temperature of the isotropic liquid phase, indeed it is preferred to cure the film some 60 degrees centigrade below the clearing point so that small local changes in temperature do not affect the birefringence of the film.
In the case of smectic phases, curing preferably takes place within the smectic phase but the constraints on temperature within the limiting temperatures of the smectic phase are not so critical as the birefringence changes only slightly with temperature in smectic phases.
The alignment process of this invention can be beneficiously applied to homeotropically align all kinds of orthogonal liquid crystals.
Preferably it can be used to align conventional monomeric non-polymerizable liquid crystals. The oriented layers then are use e.g. in the electrooptical display devices, optical compensation layers or as active switching layers e.g. in ECB or in Guest-host devices.
Most preferably according to this invention the oriented liquid crystals are reactive, polymerizable compounds, which can be either nematic or orthogonal smectic. Polymerized films of these materials are obtained by curing the layers in appropriate way and thus preserving their anisotropic orientation. These films can be removed from the substrates and also be laminated on other substrates if required. They are beneficially used to compensate optically anisotropic systems such as liquid crystal displays either alone or in combination with further anisotropic and/or isotropic films. These multilayered films are also subject of the instant invention.
Especially it is possible by the newly invented alignment method disclosed here to use one of the other layers of such a multilayered film, which typically are organic films themselves, as the substrate, due to the good alignment characteristics of the inventive alignment layers on organic substrates.

Homeotropically aligned liquid crystal layer and process for the homeotropic alignment of liquid crystals on plastic substrates